We are studying the metabolism and enzymes of 2,3-bisphosphoglycerate in the human red blood cell. It is known that 2,3-bisphosphoglycerate is important for the maintenance of the optimal oxygen carrying properties of hemoglobin. Its level changes in red cells in vivo or in vitro (e.g. stored red cells for transfusion) in response to pH and hypoxia. In various anemias the level of 2,3-bisphosphoglycerate is abnormally high. The controlling mechanisms are not yet fully understood. The 2,3-bisphosphoglycerate phosphatase of red cells is activated by a variety of anions and we are trying to define the physiologically important ones. We have found that normal human red cells contain a low concentration of the potent phosphatase activator, glycolate-2-P. Further study is underway to better define the control of the phosphatase activity. We have prepared an active site phosphohistidine peptide from a tryptic digest of red cell bisphosphoglycerate synthase. The amino acid sequence of this octapeptide is very similar to the active site peptide of yeast phosphoglycerate mutase prepared in the same way in this laboratory. These structural studies in conjunction with our kinetic work should lead to a better understanding of the mechanisms of the enzymes that use 2,3-bisphosphoglycerate.